Use of Macrolides for Treating Intestinal Inflammation

ABSTRACT

The present invention relates to a compound of formula (IA) as defined in the specification or a pharmaceutically acceptable sat thereof, in the preparation of a medicament for both the treatment of inflammatory bowel diseases and the prevention of colon cancer.

The present invention relates to novel methods of treating aninflammatory response in intestinal tissues associated with a diseasesuch as inflammatory bowel disease involving either or both the smalland large bowel. In particular the present invention relates to a newuse of in macrolides for the treatment of the inflammatory boweldisease.

Inflammatory bowel disease (IBD) is the generic term for a disease ofunknown cause that produces chronic inflammation or ulceration of themucosa of the large and small intestine. This inflammatory bowel diseaseincludes such diseases as ulcerative colitis and Crohn's disease.

Ulcerative colitis is a chronic, non-specific IBD which involves adiffuse inflammation in the mucosa of the large intestine causingulcerative lesions of the colon. Crohn's disease, also known as regionalenteritis or colitis granulomatosa, is most frequently located in thesmall intestine (small bowel), especially in the ileum, but it canaffect any part of the bowel, including the rectum. In the latter casethe differentiation of Crohn's disease from ulcerative colitis may giverise to diagnostic problems. Generally, the inflammation differs fromthat of ulcerative colitis by progressing to layers deeper than themucosa and affecting the epithelium to a lesser degree. Both diseaseshave become increasingly frequent especially in the developed countries.Therefore, treatment of IBD has become an important problem of modernmedicine.

The presently available medical treatments for IBD generally involvedrug therapy directed towards the suppression of gastrointestinalinflammation. The most commonly used medicaments to treat IBD areanti-inflammatory drugs such as the salicylates. The salicylatepreparations may be effective in treating mild to moderate disease.Examples of salicylates include sulfasalazine, olsalazine, andmesalamine. All of these medications are given orally in high doses formaximal therapeutic benefit. These medicines are not without sideeffects including heartburn, nausea, vomiting, diarrhea, and headache.

People with more severe IBD can be treated with corticosteroids, such asprednisone and hydrocortisone, which are more potent and faster-actingthan salicylates in the treatment of IBD, but are endowed withpotentially serious side effects.

In IBD patients that do not respond to salicylates or corticosteroids,medications that suppress the immune system are used. However,immunosuppressants cause increased risk of infection, renal failure, andmay increase the need for hospitalization. In severe cases of disease,the patient may need surgery to remove the affected gut. Drugs likeantidiarrheals, laxatives, and pain relievers can be also given to helprelieve symptoms.

Since all the available medical treatments for IBD are ratherunsatisfactory and often ineffective, there is currently a great needfor novel drugs capable of treating IBD and preventing its relapse.

We have now found that some macrolides, encompassed by the generalformula (I), as reported in the International patent application WO2004/013153 and endowed with anti-inflammatory activity particularly inskin and lung inflammation, are also surprisingly effective in treatingintestinal inflammation.

It is therefore a first object of the present invention the use of acompound of formula (IA)

wherein

R is hydrogen or methyl;

R¹ is an N—(C₁-C₄)acyl-N—(C₁-C₃)alkylamino group;

or a pharmaceutically acceptable salts thereof, in the preparation of amedicament for treating IBD.

Examples of pharmaceutically acceptable salts of the compounds offormula (IA) are salts with organic or mineral acids such as hydrogenchloride, hydrogen bromide, hydrogen iodine, nitric acid, sulphuricacid, phosphoric acid, acetic acid, tartaric acid, citric acid, benzoicacid, succinic acid and glutaric acid.

In particular, the compounds of formula (IA) or a pharmaceuticallyacceptable salt thereof can be used for treating ulcerative colitis orCrohn's disease.

Particularly preferred compound of formula (IA) according to theinvention is the compound of formula (ia)

namely,(9S)—O5-[3-oxoethyl)-methylamino-β-D-xylo-3,4,6-trideoxy-hexapyranosyl]-9-hydroxy-9-deoxo-erythronolide-A,or a pharmaceutically acceptable salt thereof.

Unless otherwise specified, the term “treatment” as used herein relatesto both treatment in order to cure or alleviate a disease or acondition, and to treatment in order to prevent the development orrelapse of a disease or a condition.

The term “patient”, as used herein, relates to any human or non-humanmammal in need of treatment according to the invention.

In IBD, the constant process of inflammatory injury and repair of thelining of the colon (colonic mucosa) is believed to make the individualmore susceptible to the cancer. Colon cancer doesn't distinguish betweenactive disease and remission. Patients whose disease has been quiet havethe same risk as those who have more active disease.

For example, patients with prolonged ulcerative colitis are at increasedrisk for developing colon cancer. The risk of cancer increases with theduration and the extent of involvement of colonic mucosa. For example,if only the lower colon and rectum are involved, the risk of cancer isno higher than normal. However, if the whole colon is involved, the riskof cancer may be higher.

It is therefore a further object of the present invention the use of acompound of formula (IA) as defined above or a pharmaceuticallyacceptable salt thereof for the preparation of a medicament forpreventing colon cancer.

In a further aspect, the present invention relates to a method fortreating IBD, which comprises administering to a patient in need thereofa therapeutically effective amount of a compound of formula (IA) or apharmaceutically acceptable salt thereof.

In a still further aspect the present invention relates to a method forpreventing colon cancer, which comprises administering to a patient inneed thereof a therapeutically effective amount of a compound of formula(IA) or a pharmaceutically acceptable salt thereof.

The pharmacological activity of the compound of formula (ia), as arepresentative compound of the compounds of the present invention, wasevaluated in in vivo models of intestinal inflammation, as illustratedin the following EXPERIMENTAL PART.

BRIEF DESCRIPTION OF THE DRAWINGS

In the Experimental Part below reference is made to the appendeddrawings on which:

FIG. 1 shows the scheme of drug treatments. Oral drugs were suspended in1% methocel and given in a volume of 0.5 ml.

FIG. 2 shows the macroscopic damage score criteria. The final score isobtained by the sum of all values.

FIG. 3 shows values of macroscopic damage score indicating the severityof DNBS-induced colitis in rats. Each column indicates the mean valueobtained from 6-8 animals±S.E.M. (vertical lines). Significantdifference from control values *P<0.05 (one way ANOVA following byStudent-Newman-Keuls test).

EXPERIMENTAL PART Methods Induction of Colitis

Albino male Sprague-Dawley rats, 200-250 g body weight, were treatedwith intrarectal DNBS (2,4-dinitrobenzensufonic acid, 30 mg in 0.25 mlof 50% ethanol) under a light anaesthesia with diethyl ether. In controlexperiments, the animals received 0.25 ml of saline (NaCl 0.9%). Animalswere subjected to subsequent experimental procedures 6 days afterinduction of colitis with DNBS (Blandizzi et al., Altered prejunctionalmodulation of intestinal cholinergic and noradrenergic pathways byalpha-2-adrenoceptors in the presence of experimental colitis. Br JPharmacol 139, 309-320, 2003).

Experimental Design

The compound of formula (ia), from now on the macrolide, was suspendedin 1% methocel and administered to animals by intragastric gavage in avolume of 0.5 ml at the doses of 100 or 300 μmol/kg/day for 7consecutive days. Induction of colitis was performed on day 2 asreported above. Animals were subjected to subsequent experimentalprocedures on day 7, four hours later from administration of the lastdose of the macrolide or its vehicle (FIG. 1).

Each experimental series included the following treatment groups:

-   1) controls (intragastric drug vehicle for 7 days plus intra-colonic    saline on day 2);-   2) intragastric macrolide for 7 days plus intra-colonic saline on    day 2;-   3) intragastric drug vehicle for 7 days plus intra-colonic DNBS on    day 2;-   4) intragastric macrolide for 7 days plus intra-colonic DNBS on day    2.

Each data point represents the mean value of results obtained from atleast 6-8 animals. The effect of the macrolide was compared with that ofdexamethasone (1 mg/kg/day, by oral route), known to exert a goodanti-inflammatory action in the rat model of TNBS-induced colitis(Bobin-Dubigeon et al., Effects of tumor necrosis factor-α synthesisinhibitors on rat trinitrobenzene sulfonic acid-induced chronic colitis.Eur J Pharmacol 431, 103-110, 2001). Dexamethasone was administered oncedaily following the same time schedule adopted for the macrolide.

Macroscopic and Histological Assessment of Intestinal Inflammation

At the end of treatments, animals were sacrificed and the severity ofcolonic inflammation was macroscopically evaluated in accordance withthe criteria previously reported by Wallace and Keenan (Wallace J L andKeenan C M, An orally active inhibitor of leukotriene synthesisaccelerates healing in a rat model of colitis. Am J Physiol, 258,G527-G534, 1990) with minor modifications (Blandizzi, et al., 2003).Briefly, the macroscopic evaluation is based on the following criteria:presence of adhesions between the colon and other intra-abdominalorgans; consistency of colonic faecal material (as an indirect marker ofdiarrhea); thickening of the colonic wall; presence and extension ofhyperemia and macroscopic mucosal damage (assessed with the aid of aruler) (FIG. 2). All parameters of macroscopic damage were recorded andscored for each rat by two independent observers blinded to thetreatment.

Results

The obtained result are illustrated in FIG. 3. Control rats exhibited anegligible macroscopic damage score, accounting for 1.0±0.2. InDNBS-treated animals, the extent of inflammatory lesions wassignificantly higher than that observed in control rats (10.7±0.4),indicating the occurrence of colitis. The macrolide exerted significantanti-inflammatory effects both at 100 and 300 μmol/kg (−24% and −51%,respectively). Dexamethasone induced anti-inflammatory effect similar tothat observed.with the macrolide 100 μmol/kg.

Conclusions

Data obtained from the present study indicate a good pattern ofanti-inflammatory activity of the macrolide against experimentalcolitis. In this setting, the efficacy of the macrolide seems to becomparable with that of dexamethasone, a well known anti-inflammatorydrag.

It is thus clear from the experimental evidence reported above that acompound of formula (IA), especially the compound of formula (ia) or apharmaceutically acceptable salt thereof, may be effectively used in thetreatment of IBD and in the prevention of colon cancer.

The therapeutical effective amounts of a compound of formula (IA) or apharmaceutically acceptable salt thereof will depend on the age and thegeneral physiological state of the patient, the route of administrationand the pharmaceutical formulation used; the therapeutic doses willgenerally be between about 10 and 2000 mg/day and preferably betweenabout 30 and 1500 mg/day.

The compounds of the present invention for use in the treatment and/orprophylaxis of the above mentioned diseases will preferably be used in apharmaceutical form that is suitable for oral rectal, sublingual,parenteral, topical transdermal and inhalational administration. It istherefore a further object of the present invention to providepharmaceutical formulations containing a therapeutical effective amountof a compound of formula (IA) or a pharmaceutically acceptable saltthereof together with a pharmaceutical acceptable vehicle.

The pharmaceutical formulations of the present invention may be liquid,suitable for oral and/or parenteral administration, for instance drops,syrups, solutions, injectable solutions ready to use or prepared viadilution of a lyophilizate, but preferably solid, for instance tablets,capsules, granules, powders, pellets, pessaries, suppositories, creams,pomades, gels or ointments; or alternatively solutions, suspensions,emulsions or other forms suitable for inhalation and transdermaladministration.

Depending on the type of formulation, these formulations will contain,besides a therapeutical effective amount of a compound of formula (IA)or a pharmaceutically acceptable salt thereof, solid or liquidexcipients or diluents for pharmaceutical use and optionally otheradditives normally used in the preparation of pharmaceuticalformulations, for instance thickeners, aggregating agents, lubricants,disintegrants, flavorings and colorings.

The pharmaceutical formulations according to the invention, may beproduced according to conventional techniques. As an example, hardgelatine capsules for oral administration comprising from 100 mg to 300mg of the macrolide and a pharmaceutically acceptable vehicle may beprepared to be administered to a patient in need thereof.

1. A method of treating inflammatory bowel disease, comprisingadministering an effective amount of a compound of formula (IA)

wherein R is hydrogen or methyl; R1 is anN—(C1-C4)acyl-N—(C1-C3)alkylamino group; or a pharmaceuticallyacceptable salts thereof, to a patient in need thereof.
 2. The methodaccording to claim 1, wherein the compound of formula (IA) is(9S)—O5-[3-(1-oxoethyl)-methylamino-β-D-xylo-3,4,6-trideoxy-hexapyranosyl]-9-hydroxy-9-deoxo-erythronolide-A,or a pharmaceutically acceptable salt thereof.
 3. The method accordingto claim 1, wherein the inflammatory bowel disease is ulcerativecolitis.
 4. The method according to claim 1, wherein the inflammatorybowel disease is Crohn's disease.
 5. A method, for the prevention ofcolon cancer, comprising administering an effective amount of a compoundof formula (Ia)

wherein R is hydrogen or methyl; R1 is anN—(C1-C4)acyl-N—(C1-C3)alkylamino group; or a pharmaceuticallyacceptable salts thereof, to a patient in need thereof.
 6. The methodaccording to claim 5, wherein the compound of formula (IA) is(9S)—O5-[3-(1-oxoethyl)-methylamino-β-D-xylo-3,4,6-trideoxy-hexapyranosyl]-9-hydroxy-9-deoxo-erythronolide-A,or a pharmaceutically acceptable salt thereof.
 7. 8. The method of claim1, wherein the amount of the compound of formula (IA) administered isbetween about 10 to 2,000 mg/day.
 9. The method of claim 1, wherein theamount of the compound of formula (IA) administered is between about 30to 1,500 mg/day.